Detection of CRISPR-mediated genome modifications through altered methylation patterns of CpG islands.

BACKGROUND: The development and application of CRISPR technologies for the modification of the genome are rapidly expanding. Advances in the field describe new CRISPR components that are strategically engineered to improve the precision and reliability of CRISPR editing within the genome sequence. Genome modification using induced genome breaks that are targeted and mediated by CRISPR components leverage cellular mechanisms for repair like homology directed repair (HDR) to incorporate genomic edits with increased precision. RESULTS: In this report, we describe the gain of methylation at typically hypomethylated CpG island (CGI) locations affected by the CRISPR-mediated incorporation of donor DNA using HDR mechanisms. With characterization of CpG methylation patterns using whole genome bisulfite sequencing, these CGI methylation disruptions trace the insertion of the donor DNA during the genomic edit. These insertions mediated by homology-directed recombination disrupt the generational methylation pattern stability of the edited CGI within the cells and their cellular lineage within the animal strain, persisting across generations. Our approach describes a statistically based workflow for indicating locations of modified CGIs and provides a mechanism for evaluating the directed modification of the methylome of the affected CGI at the CpG-level. CONCLUSIONS: With advances in genome modification technology comes the need to detect the level and persistence of methylation change that modifications to the genomic sequence impose upon the collaterally edited methylome. Any modification of the methylome of somatic or germline cells could have implications for gene regulation mechanisms governed by the methylation patterns of CGI regions in the application of therapeutic edits of more sensitively regulated genomic regions. The method described here locates the directed modification of the mouse epigenome that persists over generations. While this observance would require supporting molecular observations such as direct sequence changes or gene expression changes, the observation of epigenetic modification provides an indicator that intentionally directed genomic edits can lead to collateral, unintentional epigenomic changes post modification with generational persistence.

Hospitalist Co-management of Pediatric Orthopaedic Surgical Patients at a Community Hospital.

PURPOSE: The benefits of hospitalist co-management of pediatric surgical patients include bettering patient safety, decreasing negative patient outcomes, providing comprehensive medical care, and establishing a dedicated resource to patients for postoperative care. The purpose of this study was to characterize the nature of patients co-managed by a pediatric hospitalist. The authors hypothesize that hospitalist co-management is safe and efficacious in pediatric orthopaedic surgical patients who are admitted to a community hospital. METHODS: A retrospective review was performed of all pediatric orthopaedic surgical patients admitted to a community hospital who were co-managed by a pediatric hospitalist. Indications for hospitalization included pain control, antibiotic infusion, and need for neurovascular monitoring. Parameters of postoperative care and co-management were assessed, including presence of complications, medication introduction or adjustment by the hospitalist, follow-up adherence, and readmission/complication rates after discharge. RESULTS: Thirty-two patients were assessed with an average age of 8.8 years. Twenty-five percent of patients had an associated comorbidity, including asthma, attention deficit disorder, and/or autism. The pediatric hospitalist added pain medication to the original postoperative orders placed by the orthopaedics team in 44 percent of patients (14 of the 32) either for breakthrough pain or better long-term coverage. Additionally, 25 percent of patients had pain medication adjusted from the original dosing and schedule. The hospitalist team contacted the surgeon about the four patients (12.5 percent). In three of the cases, the surgeon was contacted to discuss pain medication, and one patient woke up agitated from anesthesia, necessitating a visit from the surgeon on the pediatrics floor. The length of stay was one day for all patients. The hospitalists rounded on and discharged patients the subsequent morning. All patients were given a follow-up appointment and schedule by the hospitalist team, and every patient followed up accordingly within ten days of discharge. No complications or hospital readmissions occurred within thirty days of discharge. CONCLUSION: Hospitalist co-management of pediatric orthopaedic surgical patients in a community hospital allows for better medical comorbidity and medication management. Hospitalists can provide closer observation during the inpatient stay and help streamline communication between providers and patients while allowing the surgeon the ability to be more mobile. Co-management is safe and efficacious in pediatric orthopaedic surgical patients who are admitted to a community hospital.

Hemoglobin thresholds for transfusion in pediatric patients at a large academic health center.

BACKGROUND: Although prior studies support the use of a hemoglobin (Hb) transfusion trigger of 7 to 8 g/dL for most hospitalized adults, there are few studies in pediatric populations. We therefore investigated transfusion practices and Hb triggers in hospitalized children. STUDY DESIGN AND METHODS: We performed a historical cohort study comparing transfusion practices in hospitalized children by service within a single academic institution. Blood utilization data from transfused patients (n = 3370) were obtained from electronic records over 4 years. Hb triggers and posttransfusion Hb levels were defined as the lowest and last Hb measured during hospital stay, respectively, in transfused patients. The mean and percentile distribution for Hb triggers were compared to the evidence-based restrictive transfusion threshold of 7 g/dL. RESULTS: Mean Hb triggers were above the restrictive trigger (7 g/dL) for eight of 12 pediatric services. Among all of the services, there were significant differences between the mean Hb triggers (>2.5 g/dL, p<0.0001) and between the posttransfusion Hb levels (>3 g/dL, p < 0.0001). The variation between the 10th and 90th percentiles for triggers (up to 4 g/dL, p < 0.0001) and posttransfusion Hb levels (up to 6 g/dL, p < 0.0001) were significant. Depending on the service, between 25 and 90% of transfused patients had Hb triggers higher than the restrictive range. CONCLUSIONS: Red blood cell (RBC) transfusion therapy varies significantly in hospitalized children with mean Hb triggers above a restrictive threshold for most services. Our findings suggest that transfusions may be overused and that implementing a restrictive transfusion strategy could decrease the use of RBC transfusions, thereby reducing the associated risks and costs.

Work System Assessment to Facilitate the Dissemination of a Quality Improvement Program for Optimizing Blood Culture Use: A Case Study Using a Human Factors Engineering Approach.

BACKGROUND: Work system assessments can facilitate successful implementation of quality improvement programs. Using a human factors engineering approach, we conducted a work system assessment to facilitate the dissemination of a quality improvement program for optimizing blood culture use in pediatric intensive care units at 2 hospitals. METHODS: Semistructured face-to-face interviews were conducted with clinicians from Johns Hopkins All Children's Hospital and University of Virginia Medical Center. Interview data were analyzed using qualitative content analysis. RESULTS: Blood culture-ordering practices are influenced by various work system factors, including people, tasks, tools and technologies, the physical environment, organizational conditions, and the external environment. A clinical decision-support tool could facilitate implementation by (1) standardizing blood culture-ordering practices, (2) ensuring that prescribing clinicians review the patient's condition before ordering a blood culture, (3) facilitating critical thinking, and (4) empowering nurses to communicate with physicians and advocate for adherence to blood culture-ordering guidelines. CONCLUSION: The success of interventions for optimizing blood culture use relies heavily on the local context. A work system analysis using a human factors engineering approach can identify key areas to be addressed for the successful dissemination of quality improvement interventions.

Association of a blood culture utilization intervention on antibiotic use in a pediatric intensive care unit.

Blood cultures are essential for the evaluation of sepsis. However, they may sometimes be obtained inappropriately, leading to high false-positive rates, largely due to contamination.1 As a quality improvement project, clinician decision-support tools for evaluating patients with fever or signs and symptoms of sepsis were implemented in April 2014 in our pediatric intensive care unit (PICU). This initiative resulted in a 46% decrease in blood culture obtainment2 and has been replicated in other institutions.3 It is important to evaluate antibiotic use as a balancing measure because a reduction in blood cultures could lead to an increase in antibiotic treatment days if clinicians continued empiric treatment in scenarios when blood culture results were not available. The objective of this study was to evaluate whether antibiotic use in the PICU changed in association with a reduction in blood culture utilization.

Dissemination of a Novel Framework to Improve Blood Culture Use in Pediatric Critical Care.

INTRODUCTION: Single center work demonstrated a safe reduction in unnecessary blood culture use in critically ill children. Our objective was to develop and implement a customizable quality improvement framework to reduce unnecessary blood culture testing in critically ill children across diverse clinical settings and various institutions. METHODS: Three pediatric intensive care units (14 bed medical/cardiac; 28 bed medical; 22 bed cardiac) in 2 institutions adapted and implemented a 5-part Blood Culture Improvement Framework, supported by a coordinating multidisciplinary team. Blood culture rates were compared for 24 months preimplementation to 24 months postimplementation. RESULTS: Blood culture rates decreased from 13.3, 13.5, and 11.5 cultures per 100 patient-days preimplementation to 6.4, 9.1, and 8.3 cultures per 100 patient-days postimplementation for Unit A, B, and C, respectively; a decrease of 32% (95% confidence interval, 25-43%; P < 0.001) for the 3 units combined. Postimplementation, the proportion of total blood cultures drawn from central venous catheters decreased by 51% for the 3 units combined (95% confidence interval, 29-66%; P < 0.001). Notable difference between units included the identity and involvement of the project champion, adaptions of the clinical tools, and staff monitoring and communication of project progress. Qualitative data also revealed a core set of barriers and facilitators to behavior change around pediatric intensive care unit blood culture practices. CONCLUSIONS: Three pediatric intensive units adapted a novel 5-part improvement framework and successfully reduced blood culture use in critically ill children, demonstrating that different providers and practice environments can adapt diagnostic stewardship programs.